Contraction of the heart relies upon the proper expression, trafficking, and retention of integral membrane proteins in cardiac muscle cells. These proteins vary from ion channels and transporters to hormone receptors. All play key roles in governing cardiac contraction and short and long term adaptations to physiological and pathophysiological stimuli. The profile of expressed proteins is dynamic, and it is regulated to assure the proper response to stress. This is highlighted by a decade of research linking dysfunction in membrane protein trafficking with heart disease. Yet, despite its obvious importance, little is known regarding even the identity (or the function) of the intracellular molecular pathways underpinning the trafficking and targeting of integral membrane proteins in the context of the native heart. The focus of this research program is to identify new pathways for membrane protein targeting and regulation in heart with the goal of defining novel mechanisms for the regulation of cardiac membrane excitability. Eps 15 homology domain-containing (EHD) gene products (EHD1-4) are intracellular proteins that appear to be key regulators of endosomal trafficking, lipid homeostasis, membrane protein recycling and trafficking. Previously uncharacterized in the heart, recent evidence demonstrated that this protein family likely plays indispensible roles in protein trafficking in cardiac muscle. Notably, n essential role for one of these proteins, EHD3, in the membrane trafficking of the Na/Ca exchanger (NCX) in heart was recently uncovered by our group. The goal of this proposal is to directly test the role of these proteins in cardiac structural and electrical activity using cuttin-edge in vivo models of EHD protein deficiency. Investigations will be conducted in both healthy and failing hearts. The aims of this research program are to 1) Define the in vivo roles of EHD proteins for vertebrate cardiac function at baseline and during prolonged cardiac stress, 2) identify the role of EHD proteins in the maintenance of myocyte excitability, and 3) define the role of EHD proteins in intracellular protein trafficking. The work completed here will be the firs to characterize the functional molecular mechanisms underpinning both protein trafficking and electrical remodeling within the heart.